Synaptic plasticity is accompanied by a change in the protein composition of synapses. Recent published findings support the idea that regulated proteolysis via the ubiquitin proteasome system (UPS) is required to promote, limit or restrict modulations in synaptic strength. In mammals, proteasome inhibition alters basal synaptic transmission, long-term potentiation (LTP), long-term depression (LTD) and behavior. The proposed work is designed to test the validity and functional relevance of proteasome phosphorylation as a cis-related regulatory mechanism to control its activity in neurons. The reasoning behind this line of investigation is a direct result of our recent findings. In short, we find the following: proteasome function is directly modified by increased and decreased neuronal activity;proteasomes are phosphorylated in neurons in an activity dependent fashion;CaMKII, a key plasticity kinase, regulates proteasome activity;and CaMKII directly phosphorylates the 19S regulatory cap of the 26S proteasome. Our data therefore uncover a novel biochemical signaling pathway linking neuronal activity, a key plasticity kinase (CaMKII), and the dynamic regulation of proteasome function. There are two main goals of this proposal. Firstly, we will determine how phosphorylation by CaMKII affects the activity and distribution of proteasomes in neurons (AIM1). Secondly, we will determine if the phosphorylation of the proteasome is involved in synaptic function (AIM2). Sites of phosphorylation will first be identified, followed by several experimental strategies to determine how altered proteasome phosphorylation affects its activity and distribution in neurons under control and activity manipulated conditions. The functional relevance for synaptic plasticity will then be explored. We predict neurons to utilize this mechanism to synergize the dynamic targeting of proteasome substrates with that of tunable proteasome activity to facilitate modulations in synaptic strength. PUBLIC HEALTH RELEVANCE: Synaptic plasticity is accompanied by a change in the protein composition of synapses. The ubiquitin proteasome system (UPS) is one of the major cellular pathways which controls protein turnover in eukaryotic cells. While the UPS has been widely attributed to and extensively studied in neurodegenerative disease, the function of the UPS in normal neuronal physiology, however, is far less understood. This proposal explores the synaptic function of the UPS in the mammalian central nervous system (CNS). We have discovered that neuronal activity and phosphorylation by a key plasticity kinase, CaMKII, are involved in the dynamic regulation of proteasome function in neurons. The proposed work is designed to test the validity and functional relevance of proteasome phosphorylation as a cis-related regulatory mechanism to control its activity in neurons. These studies will be important for understanding the role of the UPS in synaptic function as well as synaptic dysfunction often associated with neurodegenerative disease.